Objectives To test the hypothesis that somatic PIK3CA mutations would be found in patients with more common disorders including isolated lymphatic malformation (LM) and Klippel-Trenaunay syndrome (KTS). Study design We used next generation sequencing, droplet digital PCR (ddPCR), and single molecule molecular inversion probes (smMIPs) to search for somatic PIK3CA mutations in affected tissue from patients seen at Boston Children’s Hospital who had an isolated LM (n=17), KTS (n=21), fibro-adipose vascular anomaly (FAVA; n=8), or congenital lipomatous overgrowth with vascular, epidermal, and skeletal anomalies syndrome (CLOVES; n = 33), the disorder for which we first identified somatic PIK3CA mutations. We also screened 5 of the more common PIK3CA mutations in a second cohort of patients with LM (n=31) from Seattle Children’s Hospital. Results Most individuals from Boston Children’s Hospital who had isolated LM (16/17) or LM as part of a syndrome, such as KTS (19/21), FAVA (4/8), and CLOVES (30/32) were somatic mosaic for PIK3CA mutations, with 5 specific PIK3CA mutations accounting for ~ 80% of cases. Seventy-four percent of patients with LM from Seattle Children’s Hospital also were somatic mosaic for 1 of 5 specific PIK3CA mutations. Many affected tissue specimens from both cohorts contained fewer than 10% mutant cells. Conclusions Somatic PIK3CA mutations are the most common cause of isolated lymphatic malformations and disorders in which lymphatic malformation is a component feature. Five PIK3CA mutations account for most cases. The search for causal mutations requires sampling of affected tissues and techniques that are capable of detecting low-level somatic mosaicism, because the abundance of mutant cells in a malformed tissue can be low.

Background A somatic mutation in GNAQ (c.548G>A;p.R183Q), encoding Gαq, has been found in syndromic and sporadic capillary malformation tissue. However, the specific cell type(s) containing the mutation is unknown. The purpose of this study was to determine which cell(s) in capillary malformations have the GNAQ mutation. Methods Human capillary malformation tissue was obtained from 13 patients during a clinically-indicated procedure. Droplet digital PCR (ddPCR), capable of detecting mutant allelic frequencies as low as 0.1%, was used to quantify the abundance of GNAQ mutant cells in capillary malformation tissue. Six specimens were fractionated by fluorescence activated cell sorting (FACS) into hematopoietic, endothelial, perivascular, and stromal cells. The frequency of GNAQ mutant cells in these populations was quantified by ddPCR. Results Eight capillary malformations contained GNAQ p.R183Q mutant cells, 2 lesions had novel GNAQ mutations (p.R183L; p.R183G), and 3 capillary malformations did not have a detectable GNAQ p.R183 mutation. Mutant allelic frequencies ranged from 2% to 11%. Following FACS, the GNAQ mutation was found in the endothelial but not the platelet-derived growth factor receptor-β-positive (PDGFRβ) cell population; mutant allelic frequencies were 3% to 43%. Conculsions Endothelial cells in capillary malformations are enriched for GNAQ mutations and are likely responsible for the pathophysiology underlying capillary malformation.